Drug resistance has become a major challenge in cancer treatment, but the mechanisms are just being discovered. Emerging evidence suggest that some drug resistance mechanisms are due to epigenetic changes and can be targeted to overcome drug resistance. Despite the development of modern genomic and epigenomic methods, there is no systematic approach to discriminate the driver and passenger epigenetic events. Thus, identification of the driver epigenetic changes that cause drug resistance with a systematic approach is essential for the development of cancer therapeutic strategies to target these changes. The long-term goal is to elucidate the roles of epigenetic events that drive tumor initiation and progression and to translate these findings to the clinic. The objective of this proposal is to identify the critical epigenetic events that confer trastuzuma resistance of HER2+ breast cancer cells. The central hypothesis is that epigenetic changes play a major role in trastuzumab resistance. The hypothesis is based mainly on the preliminary data produced with a unique cell-based system on trastuzumab resistance. The rationale for the proposed research is that better understanding of epigenetic driver events for trastuzumab resistance will result in new and innovative approaches to breast cancer treatment. Therefore, the hypothesis will be tested by pursuing the following two specific aims: 1) Identify the epigenetic regulators that are necessary for trastuzumab resistance, 2) Determine the roles of the epigenetic changes caused by the identified epigenetic regulators in trastuzumab resistance. The proposed research is innovative, because of the novel hypothesis that a major trastuzumab resistance mechanism is epigenetic changes and the systematic approach to integrate a functional screen with targeted epigenomic analysis of the unique cell based system. The proposed research is significant, because it is expected to vertically advance and expand our understanding of epigenetic regulation in drug resistance. Such knowledge is critical for the development of cancer therapies targeting epigenetic aberrations.